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Preserving the extra cellular matrix is critical for improving efficacy of keratinocyte sheets for treatment of burn wounds
Author(s) -
Alharbi Suzan,
Niimi Yosuke,
Popov Vsevolod,
Hawkins Hal,
Cox Robert,
Wiener Dominique,
Osada Atsuyoshi,
Ihara Koji,
Sakurai Hiroyuki,
Herndon David,
Enkhbaatar Perenlei
Publication year - 2019
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.2019.33.1_supplement.123.1
Subject(s) - keratinocyte , epidermis (zoology) , burn wound , medicine , wound healing , skin grafting , cadaver , organ culture , surgery , dermatology , chemistry , anatomy , in vitro , biochemistry
Cultured autologous keratinocyte sheets (KS) have been proposed for treating burn wounds for over 30 years; however, they have not been translated to clinical practice because of negative effects of enzymes, used for detachment, on KS quality. The aim of this study was to test the hypothesis that non‐enzymatically detached KS exert better effects on grafted burn wound healing compared to enzymatically detached KS. Methods For non‐enzymatic detachment, autologous ovine primary keratinocytes were cultured in novel temperature‐responsive dishes (T sheets), and were detached by temperature reduction to 20°C. For comparison, the aliquots of keratinocytes were also cultured in conventional dish to be detached by use of enzyme (Dispase) (D sheets). We have tested the efficacy of these sheets using a well‐characterized, novel and clinically relevant ovine model of grafted burn wounds. Briefly, six full thickness skin burns (5×5cm) were induced in sheep dorsum under anesthesia and analgesia. After 24 hrs, burned skin was excised and wounds were grafted with ovine cadaver skin. After 3 weeks, rejected cadaver skin epidermis was debrided and wounds were covered with T and D sheets. Thereafter, the wounds were assessed for two more weeks. Wound samples were collected postmortem for various assays. The autologous keratinocytes for the sheet culture were isolated at the same day of skin grafting which mimics the clinical situation. Results Cultured keratinocytes in both dishes formed firm sheets within three weeks, mimicking the time of cadaver skin epidermis rejection. Keratinocyte sheet thickness was significantly higher in T sheets compared to D sheets. In vivo , T sheets were better accepted with significantly higher wound re‐epithelialization percentage than D sheets at days 7 and 14 after sheet grafting. Although the epidermis thickness was comparable in wounds treated with both types of sheets, the dermal‐epidermal junction was well defined in wounds covered with T sheets at 14 th day as evidenced by continuous lamina densa and high number of hemi‐desmosomes. Histopathological evaluation for burn wounds revealed that wounds treated with T sheets had less hemorrhage, ulceration incidence and neutrophils infiltration. Conclusion Novel non‐enzymatic method for KS detachment provides better sheet quality and accelerates healing of grafted burn wounds. Support or Funding Information Shriner's Hospital for Children SHC85100 and SHC84050 This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .